Regeneration-Associated Cells Improve Recovery from Myocardial Infarction through Enhanced Vasculogenesis, Anti-inflammation, and Cardiomyogenesis

Abstract

ABSTRACTBackgroundConsidering the impaired function of regenerative cells in patients with comorbidities and associated risk factors, cell therapy to enhance the regenerative microenvironment was designed using regeneration-associated cells (RACs), including endothelial progenitor cells (EPCs) and anti-inflammatory cells.MethodsRACs were prepared by quality and quantity control culture of blood mononuclear cells (QQMNCs). The peripheral blood mononuclear cells (PBMNCs) were isolated from Lewis rats and conditioned for 5 days using a medium containing stem cell factors, thrombopoietin, Flt-3 ligand, vascular endothelial growth factor, and interleukin-6 to generate QQMNCs.ResultsIn vitro EPC colony forming assays demonstrated a 5.3-fold increase in the definitive colony-forming EPCs and vasculogenic EPCs, in comparison to naïve PBMNCs. Flow cytometry analysis revealed that QQMNCs were enriched with RACs, such as EPCs (28.9-fold, P<0.0019) and M2 macrophages (160.3-fold, P<0.0002). Cell transcriptome analysis revealed that angiogenesis (angpt1, angpt2, and vegfb), stem/progenitor (c-kit and sca-1) and anti-inflammation related (arg-1, erg-2, tgfb, and foxp3) genes were highly expressed in QQMNCs. For in vivo cell transplantation experiments, 1×105 cells were administered via the tail vein into syngeneic rat models of myocardial infarction (MI). Echocardiographic data showed that QQMNCs-transplanted group (QQ-Tx) preserved cardiac function and fraction shortening (45.5±4.6%) at 28-days post-MI in comparison with PBMNCs-transplanted (PB-Tx) (30.9±6.4%, P<0.0001) and Control (32.2±7.7%, P<0.0008) groups. Histological analysis revealed that QQ-Tx showed enhanced angiogenesis and reduced interstitial left ventricular fibrosis, along with a decrease in neutrophils and an increase in M2 macrophages in the acute phase of MI. Cell tracing studies revealed that intravenously administered QQMNCs preferentially homed to ischemic tissues via blood circulation, while PBMNCs did not. QQ-Tx showed markedly upregulated early cardiac transcriptional cofactors (Nkx2-5, 29.8-fold, and Gata-4, 5.2- fold) as well as c-kit (4.5-fold) while these markers were downregulated in PB-Tx. In QQ-Tx animals, de novo blood vessels formed a “Biological Bypass” as observed macroscopically and microscopically, while PB-Tx and Control-Tx groups never developed epicardial blood vessels but showed severe fibrotic adhesion to the surrounding tissues.ConclusionQQMNCs, as RACs derived from rat PBMNCs, conferred potent angiogenic and anti-inflammatory properties to the regenerative microenvironment, enhancing myocardiogenesis and functional recovery of rat MI hearts.